Cardiovascular disease is a major health risk throughout the industrialized world. Atherosclerosis, the most prevalent of cardiovascular diseases, is the principal cause of heart attack, stroke, and peripheral vascular disease resulting in significant disability and limb loss, and thereby the principle cause of death in the United States.
Atherosclerosis is a complex disease involving many cell types and molecular factors (described in, for example, Ross (1993) Nature 362: 801–809). The process, in normal circumstances a protective response to insults to the endothelium and smooth muscle cells (SMCs) of the wall of the artery, consists of the formation of fibrofatty and fibrous lesions or plaques, preceded and accompanied by inflammation. The advanced lesions of atherosclerosis may occlude the artery concerned, and result from an excessive inflammatory-fibroproliferative response to numerous different forms of insult. Injury or dysfunction of the vascular endothelium is a common feature of many conditions that predispose an individual to accelerated development of atherosclerotic cardiovascular disease. For example, shear stresses are thought to be responsible for the frequent occurrence of atherosclerotic plaques in regions of the circulatory system where turbulent blood flow occurs, such as branch points and irregular structures.
The first observable event in the formation of an atherosclerotic plaque occurs when blood-borne monocytes adhere to the vascular endothelial layer and transmigrate through to the sub-endothelial space. Adjacent endothelial cells at the same time produce oxidized low density lipoprotein (LDL). These oxidized LDLs are then taken up in large amounts by the monocytes through scavenger receptors expressed on their surfaces. In contrast to the regulated pathway by which native LDL (nLDL) is taken up by nLDL specific receptors, the scavenger pathway of uptake is not regulated by the monocytes.
These lipid-filled monocytes are called foam cells, and are the major constituent of the fatty streak. Interactions between foam cells and the endothelial and smooth muscle cells which surround them lead to a state of chronic local inflammation which can eventually lead to smooth muscle cell proliferation and migration, and the formation of a fibrous plaque.
Such plaque may totally or partially block blood flow through a blood vessel leading to a heart attack or stroke. Plaque can also weaken the arterial wall, resulting in an aneurysm. Moreover, occlusion of the blood vessels caused by plaques restrict the flow of blood, resulting in ischemia. Ischemia is a condition characterized by a lack of oxygen supply in tissues of organs due to inadequate perfusion. Such inadequate perfusion can have a number of natural causes, including atherosclerotic or restenotic lesions, anemia, or stroke. Many medical interventions, such as the interruption of the flow of blood during bypass surgery, for example, also lead to ischemia. In addition to sometimes being caused by diseased cardiovascular tissue, ischemia may sometimes affect cardiovascular tissue, such as in ischemic heart disease. Ischemia may occur in any organ, however, that is suffering a lack of oxygen supply.
The P2X receptors are a family of ligand-gated membrane ion channels activated by the binding of extracellular adenosine 5′-triphosphate (ATP). Seven different P2X receptor subunit cDNAs have been identified (P2X1, P2X2, P2X3, P2X4, P2X5, P2X6, and P2X7) (MacKenzie, et al. (1999) Ann. N.Y. Acad. Sci. 868:716–729). They are characterized by two transmembrane domains with a large extracellular loop where 10 cysteine residues are preserved; and by intracellular N- and C-terminals (Burnstock (2000) British Journal of Anesthesia 84:476–880). P2X receptors are widely distributed in various tissues of mammals, including smooth muscle of the urinary bladder and arteries, kidney, pancreas, lung, cardiac myocytes, sensory and sympathetic ganglia, brain and spinal cord, and each subtype seems to be preferentially expressed in different tissue (Yamamoto, et al. (2000) Am. J. Pilysiol. Heart Circ. Physiol. 279:H285–H292).
The human P2X4 gene was cloned from the brain and forms functional homomeric ATP-activated channels when expressed in heterologous cellular systems (Garcia-Guzman, et al. (1997) Molecular Pharmacology 51:109–118). This receptor has been found to be expressed in human endothelial cells, and is involved in ATP-induced Ca2+ influx in endothelial cells (Yamamoto, et al. (2000) Am. J. Physiol. Heart Circ. Physiol. 279:H285–H292).
Calcium concentration plays a role in cardiovascular diseases, including atherosclerosis. Calcium channel blockers (CCB) have been used to effectively modulate high blood pressure. It has been postulated that CCB's could also be used to avoid calcium deposits in arterial walls, which is one of the main components of atherosclerotic plaques (Perez (2000) J. Hum. Hypertens. 14 Suppl 1:S96–9). Intracelllular calcium levels have also been correlated with late phase platelet aggregation and formation of a hemostaic plug, which has been implicated in the pathogenesis of atherosclerosis (Covic, et al. (2000) Biochemistry 39:5458–5467). Recent studies also have focused on the role of free radicals on calcium signaling. Vascular calcium signaling is altered by oxidant stress in ischemia-related disease states (Lounsbury et al. (2000) Free Radical. Biol. Med. 28:1362). Extracellular calcium has been shown to function as an ionic chemokinetic agent capable of modulating the innate immune response in vivo and in vitro by direct and indirect actions on monocytic cells. Therefore, calcium deposition may be both a consequence of and/or a cause of chronic inflammatory changes at sites of injury, infection, and atherosclerosis (Olszak, et al. (2000) J. Clin. Invest. 105:1299–305).